Pharmaceutical compositions comprising perillyl alcohol derivatives

ABSTRACT

A pharmaceutical composition is provided which includes perillyl alcohol conjugated with a therapeutic agent and further includes and a hydrolyzable acylated aliphatic tail. A method of using the pharmaceutical composition is also provided for treating a condition or disease of a patient, e.g., cancer.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Patent ApplicationSer. No. 62/115,396, filed on Feb. 12, 2015 which is incorporated byreference in its entirety.

FIELD OF THE INVENTION

The present invention relates to monoterpene derivatives containing atherapeutic agent as well as an aliphatic tail. The present inventionfurther relates to methods and compositions of using the monoterpenederivatives to treat cancer.

BACKGROUND

Malignant gliomas, the most common form of central nervous system (CNS)cancers, is currently considered essentially incurable. Among thevarious malignant gliomas, anaplastic astrocytomas (Grade III) andglioblastoma multiforme (GBM; Grade IV) have an especially poorprognosis due to their aggressive growth and resistance to currentlyavailable therapies. The present standard of care for malignant gliomasconsists of surgery, ionizing radiation, and chemotherapy. Despiterecent advances in medicine, the past 50 years have not seen anysignificant improvement in prognosis for malignant gliomas. Wen et al.Malignant gliomas in adults. New England J Med. 359: 492-507, 2008.Stupp et al. Radiotherapy plus concomitant and adjuvant temozolomide forglioblastoma. New England J Med. 352: 987-996, 2005.

The poor response of tumors, including malignant gliomas, to varioustypes of chemotherapeutic agents are often due to intrinsic drugresistance. Additionally, acquired resistance of initiallywell-responding tumors and unwanted side effects are other problems thatfrequently thwart long-term treatment using chemotherapeutic agents.Hence, various analogues of chemotherapeutic agents have been preparedin an effort to overcome these problems. The analogues include noveltherapeutic agents which are hybrid molecules of at least two existingtherapeutic agents. For example, cisplatin has been conjugated withPt-(II) complexes with cytotoxic codrugs, or conjugated with bioactiveshuttle components such as porphyrins, bile acids, hormones, ormodulators that expedite the transmembrane transport or the drugaccumulation within the cell. (6-Aminomethylnicotinate)dichloridoplatinum(II) complexes esterified with terpene alcohols weretested on a panel of human tumor cell lines. The terpenyl moieties inthese complexes appeared to fulfill a transmembrane shuttle function andincreased the rate and extent of the uptake of these conjugates intovarious tumor cell lines. Schobert et al. Monoterpenes as Drug Shuttles:Cytotoxic (6-minomethylnicotinate) dichloridoplatinum(II) Complexes withPotential to Overcome Cisplatin Resistance. J. Med. Chem. 2007, 50,1288-1293.

Perillyl alcohol (POH), a naturally occurring monoterpene, has beensuggested to be an effective agent against a variety of cancers,including CNS cancer, breast cancer, pancreatic cancer, lung cancer,melanomas and colon cancer. Gould, M. Cancer chemoprevention and therapyby monoterpenes. Environ Health Perspect. 1997 June; 105 (Suppl 4):977-979. Hybrid molecules containing both perillyl alcohol and retinoidswere prepared to increase apoptosis-inducing activity. Das et al. Designand synthesis of potential new apoptosis agents: hybrid compoundscontaining perillyl alcohol and new constrained retinoids. TetrahedronLetters 2010, 51, 1462-1466.

Co-owned U.S. Patent Publication No. 20130203828 disclosed variousperillyl alcohol derivatives such as perillyl alcohol carbamates. Forexample, the perillyl alcohol derivatives include perillyl alcoholconjugated with a therapeutic agent such as dimethyl celocoxib (DMC),temozolomide (TMZ) or rolipram.

There is still a need for perillyl alcohol derivatives with improvedproperties for transdermal or topical application, as well as use thederivatives in the treatment of cancers such as malignant gliomas, skincancers as well as precancerous skin conditions.

SUMMARY OF THE INVENTION

In one aspect of the present application, a pharmaceutical compositionis provided. The composition comprises a compound comprising perillylalcohol conjugated with a therapeutic agent, and an acylated aliphatictail. The aliphatic tail can be derived from fatty acids. In someembodiments, the aliphatic tail contains 4 to 28 carbon atoms. Thealiphatic chain can be saturated or unsaturated, branched ornon-branched.

In one embodiment, the compound is a perillyl alcohol carbamate, wherethe nitrogen of the carbamate group is acylated with the aliphatic tail.In one embodiment, the therapeutic agent is a chemotherapeutic agent.The chemotherapeutic agent includes, but is not limited to, of a DNAalkylating agent, a topoisomerase inhibitor, an endoplasmic reticulumstress inducing agent, a platinum compound, an antimetabolite, an enzymeinhibitor, and a receptor antagonist. In specific embodiments, thetherapeutic agent can be dimethyl celocoxib (DMC), temozolomide (TMZ),or rolipram.

In one embodiment, the compound is(3-Methyl-4-oxo-3,4-dihydro-imidazo[5,1-d][1,2,3,5]tetrazine-8-carbonyl)-octadeca-9,12-dienoyl-carbamicacid 4-isopropenyl-cyclohex-1-enylmethyl ester. In another embodiment,the compound is(3-Methyl-4-oxo-3,4-dihydro-imidazo[5,1-d][1,2,3,5]tetrazine-8-carbonyl)-hexadecanoicacid 4-isopropenyl-cyclohex-1-enylmethyl ester.

In another aspect, there is provided compounds of the Formula I;

wherein R is selected from a C₄ to C₂₈ linear or branched alkyl, C₄ toC₂₈ alkeneyl or a C₄ to C₂₈ alkynyl group; or a pharmaceuticallyacceptable salt thereof.

In another aspect there is provided a compound according to Formula Iwherein R is a C₄ to C₂₈ linear or branched alkeneyl containing 1, 2, 3,4 or 5 double bonds.

In another aspect there is provided a compound according to Formula Iwherein R is a C₄ to C₂₈ linear or branched alkynyl containing 1, 2, 3,4 or 5 triple bonds.

In another aspect a compound of the Formula II;

or a pharmaceutically acceptable salt thereof is provided.In another aspect, a compound of the Formula III is provided;

wherein R₁ is selected from a C₄ to C₂₈ linear or branched alkyl, linearor branched C₄ to C₂₈ alkeneyl or a linear or branched C₄ to C₂₈ alkynylgroup; or a pharmaceutically acceptable salt thereof.In another aspect there is provided a compound according to Formula IIIwherein R₁ is a C₄ to C₂₈ linear or branched alkeneyl containing 1, 2,3, 4 or 5 double bonds.

In another aspect there is provided a compound according to Formula IIIwherein R₁ is a C₄ to C₂₈ linear or branched alkynyl containing 1, 2, 3,4 or 5 triple bonds.

In another aspect a compound of the Formula IV; or a pharmaceuticallyacceptable salt thereof is provided.

In another aspect, the present invention provides a method for treatinga disease in a mammal. The method comprises delivering to the mammal atherapeutically effective amount of a pharmaceutical compositiondescribed herein. In one embodiment, the disease is cancer. For example,the cancer is a tumor of the nervous system, e.g., glioblastoma. Thecancer can also be a skin cancer, such as melanoma, basal cellcarcinoma, and squamous cell carcinoma. In another embodiment, thedisease is a precancerous skin lesion. The composition can beadministered by inhalation, intranasally, orally, intravenously,topically, transdermally, subcutaneously or intramuscularly.

In another aspect a process for making a compound of the Formula I

comprising the steps of

a) reacting Temozolamide with oxalyl chloride in a halogenated solvent,preferably 1,2 dichloroethane, to give the isocyanate of TemozolamideFormula A

b) reacting the isocyanate of Temozolamide with perillyl alcohol in ahalogenated solvent to afford a compound of Formula B

c) reacting the compound of Formula B with an acid chloride, preferablylinoleic acid chloride, of the Formula C

wherein R is selected from the group consisting of a C₄ to C₂₈ linear orbranched alkyl, linear or branched C₄ to C₂₈ alkeneyl containing 1, 2,3, 4, or 5 double bonds or a linear or branched C₄ to C₂₈ alkynyl group;containing 1, 2, 3, 4 or 5 triple bonds in the presence of a base,preferably NaH, in an ethereal solvent, preferably THF, to afford acompound of the Formula I is provided.

BRIEF DESCRIPTION OF THE DRAWINGS

The patent or application file contains at least one drawing executed incolor. Copies of this patent or patent application publication withcolor drawing(s) will be provided by the Office upon request and paymentof the necessary fee.

FIG. 1 is a plot showing in vitro efficacy of temozolomide (TMZ),TMZ-POH, and TMZ-POH-linoleate (TPL) in the treatment of MGMT negativehuman melanoma cells A2058.

FIG. 2 is a plot showing in vitro efficacy of temozolomide (TMZ),TMZ-POH, and TPL in the treatment of MGMT positive human melanoma cellsA375.

FIG. 3A is an image of a tumor formed on the skin of a nude mouse due tosubcutaneously injected melanoma cells.

FIG. 3B is an image of the tumor depicted in FIG. 3A after being treatedwith TPL for 14 days.

FIGS. 4A-4C are example micrographs of the tissue stained forvisualization of CD31 marker expression obtained from a tumor formed onthe skin of a nude mouse due to subcutaneously injected melanoma cells,where the mouse had been treated by a vehicle for 14 days.

FIGS. 4D-4F are example micrographs of the tissue stained forvisualization of CD31 marker expression obtained from a nodule formed onthe skin of a nude mouse due to subcutaneously injected melanoma cells,where the mouse had been treated by TPL for 14 days.

FIG. 5 is a plot comparing the average area representing CD31 markerexpression in a nodule formed on the skin of a nude mouse due tosubcutaneously injected melanoma cells which had been treated with TPLfor 14 days with that of a control mouse treated with a vehicle.

DETAILED DESCRIPTION

In one aspect, the present invention provides for derivatives ofmonoterpene (or sesquiterpene) comprising a monoterpene (orsesquiterpene) conjugated with a therapeutic agent. The monoterpene (orsesquiterpene) derivative can further include an acylated aliphatictail. In some embodiments, the aliphatic tail contains 4 to 28 carbonatoms. As an example, the monoterpene (or sesquiterpene) can be perillylalcohol (POH). The therapeutic agent may be covalently linked with themonoterpene (or sesquiterpene) through carbamate, ester, ether bonds, orany other suitable chemical functional groups. For example, themonoterpene derivative can be perillyl alcohol carbamate of atherapeutic agent wherein the nitrogen of the carbamate group isacylated with the aliphatic tail. The therapeutic agent can be achemotherapeutic agent, such as a DNA alkylating agent, a topoisomeraseinhibitor, an endoplasmic reticulum stress inducing agent, a platinumcompound, an antimetabolite, an enzyme inhibitor, or a receptorantagonist. In particular embodiments, the therapeutic agent can bedimethyl celocoxib (DMC), temozolomide (TMZ), or rolipram. The molarratio of the monoterpene (or sesquiterpene) to the therapeutic agent inthe monoterpene (or sesquiterpene) conjugate may be 1:1, 1:2, 1:3, 1:4,2:1, 3:1, 4:1, or any other suitable molar ratios. When the monoterpene(or sesquiterpene) and the therapeutic agent are conjugated through acarbamate bond, the therapeutic agent may be any agent bearing at leastone carboxylic acid functional group, or any agent bearing at least oneamine functional group. The aliphatic chain can be saturated orunsaturated, straight chain or branched-chain.

As used herein, monoterpenes include terpenes that consist of twoisoprene units. Monoterpenes may be linear (acyclic) or contain rings.Derivatives of monoterpenoids are also encompassed by the presentinvention. Monoterpenoids may be produced by biochemical modificationssuch as oxidation or rearrangement of monoterpenes. Examples ofmonoterpenes and monoterpenoids include, perillyl alcohol (S(−)) and(R(+)), ocimene, myrcene, geraniol, citral, citronellol, citronellal,linalool, pinene, terpineol, terpinen, limonene, terpinenes,phellandrenes, terpinolene, terpinen-4-ol (or tea tree oil), pinene,terpineol, terpinen; the terpenoids such as p-cymene which is derivedfrom monocyclic terpenes such as menthol, thymol and carvacrol; bicyclicmonoterpenoids such as camphor, borneol and eucalyptol.

Monoterpenes may be distinguished by the structure of a carbon skeletonand may be grouped into acyclic monoterpenes (e.g., myrcene, (Z)- and(E)-ocimene, linalool, geraniol, nerol, citronellol, myrcenol, geranial,citral a, neral, citral b, citronellal, etc.), monocyclic monoterpenes(e.g., limonene, terpinene, phellandrene, terpinolene, menthol, carveol,etc.), bicyclic monoterpenes (e.g., pinene, myrtenol, myrtenal,verbanol, verbanon, pinocarveol, carene, sabinene, camphene, thujene,etc.) and tricyclic monoterpenes (e.g. tricyclene). See Encyclopedia ofChemical Technology, Fourth Edition, Volume 23, page 834-835.

Sesquiterpenes of the present invention include terpenes that consist ofthree isoprene units. Sesquiterpenes may be linear (acyclic) or containrings. Derivatives of sesquiterpenoids are also encompassed by thepresent invention. Sesquiterpenoids may be produced by biochemicalmodifications such as oxidation or rearrangement of sesquiterpenes.Examples of sesquiterpenes include farnesol, farnesal, farnesylic acidand nerolidol.

The derivatives of monoterpene (or sesquiterpene) include, but are notlimited to, carbamates, esters, ethers, alcohols and aldehydes of themonoterpene (or sesquiterpene). Monoterpene (or sesquiterpene) alcoholsmay be derivatized to carbamates, esters, ethers, aldehydes or acids.

Carbamate refers to a class of chemical compounds sharing the functionalgroup

based on a carbonyl group flanked by an oxygen and a nitrogen. The Rgroups on the nitrogen and the oxygen may form a ring. R¹—OH may be amonoterpene, e.g., POH. In some embodiments, one of R² and R³ may be atherapeutic agent, while the other of R² and R³ is an acylated aliphaticchain having 4-28 carbons.

Carbamates may be synthesized by reacting isocyanate and alcohol, or byreacting chloroformate with amine. Carbamates may be synthesized byreactions making use of phosgene or phosgene equivalents. For example,carbamates may be synthesized by reacting phosgene gas, diphosgene or asolid phosgene precursor such as triphosgene with two amines or an amineand an alcohol. Carbamates (also known as urethanes) can also be madefrom reaction of a urea intermediate with an alcohol. Dimethyl carbonateand diphenyl carbonate are also used for making carbamates.Alternatively, carbamates may be synthesized through the reaction ofalcohol and/or amine precursors with an ester-substituted diarylcarbonate, such as bismethylsalicylcarbonate (BMSC). U.S. PatentPublication No. 20100113819.

Carbamates may be synthesized by the following approach:

Monoterpenes or sesquiterpines (e.g POH) are reacted with phosgene inthe presence of a first base such and an aromatic solvent such astoluene to form the corresponding chloroformate. The chloroformates arethen reacted with a therapeutic agent having an NH₂ group (e.g DMC)optionally in the presence of a second base to afford the carbamate. Thecarbamates can then be reacted with an acid chloride of a C₄ to C₂₈linear or branched alkyl carboxylic acid, a C₄ to C₂₈ alkeneylcarboxylic acid having 1-5 double bonds or a C₄ to C₂₈ alkynylcarboxylic acid having 1-5 triple bonds; optionally in the presence of athird base. Suitable reaction solvents include, but are not limited to,tetrahydrofuran, dichloromethane, dichloroethane, acetone, anddiisopropyl ether. The reaction may be performed at a temperatureranging from about −70° C. to about 80° C., or from about −65° C. toabout 50° C. The molar ratio of perillyl chloroformate (or thechloroformate of a monoterpene or sesquiterpene) to the substrate R₁—NH₂(wherein R₁—NH₂ is a therapeutic agent) may range from about 1:1 toabout 2:1, from about 1:1 to about 1.5:1, from about 2:1 to about 1:1,or from about 1.05:1 to about 1.1:1. Suitable first, second and thirdbases include, but are not limited to, organic bases, such astriethylamine, N,N′-diisopropylethylamine, butyl lithium, andpotassium-t-butoxide and inorganic bases such as sodium or potassiumcarbonate, KOH, NaOH and NaH.

Alternatively, carbamates may be synthesized by the following approach:

R₂C(O)NH₂ (where R₂C(O)NH₂ is a therapeutic agent e.g TMZ) is reactedwith oxalyl chloride to produce the isocyanate (R₂—N═C═O) followed byreaction with a monoterpene or sesquiterpene (e.g POH) to afford thecarbamate, optionally in the presence of a first base. The carbamatescan then be reacted with an acid chloride of a C₄ to C₂₈ linear orbranched alkyl carboxylic acid, a C₄ to C₂₈ alkeneyl carboxylic acidhaving 1-5 double bonds or a C₄ to C₂₈ alkynyl carboxylic acid having1-5 triple bonds; optionally in the presence of a second base. Suitablereaction solvents include, but are not limited to, dichloromethane,dichloroethane, toluene, diisopropyl ether, and tetrahydrofuran. Thereaction may be performed at a temperature ranging from about 25° C. toabout 110° C., or from about 30° C. to about 80° C., or about 50° C. Themolar ratio of perillyl alcohol (or monoterpene or sesquiterpene) to thesubstrate R₂—N═C═O may range from about 1:1 to about 2:1, from about 1:1to about 1.5:1, from about 2:1 to about 1:1, or from about 1.05:1 toabout 1.1:1. Suitable first, second and third bases include, but are notlimited to, organic bases, such as triethylamine,N,N′-diisopropylethylamine, butyl lithium, and potassium-t-butoxide orinorganic bases such as but not limited to potassium or sodiumcarbonate, NaOH, KOH, and NaH.

Esters of the monoterpene (or sesquiterpene) alcohols of the presentinvention can be derived from an inorganic acid or an organic acid.Inorganic acids include, but are not limited to, phosphoric acid,sulfuric acid, and nitric acid. Organic acids include, but are notlimited to, carboxylic acid such as benzoic acid, fatty acid, aceticacid and propionic acid, and any therapeutic agent bearing at least onecarboxylic acid functional group Examples of esters of monoterpene (orsesquiterpene) alcohols include, but are not limited to, carboxylic acidesters (such as benzoate esters, fatty acid esters (e.g., palmitateester, linoleate ester, stearate ester, butyryl ester and oleate ester),acetates, propionates (or propanoates), and formates), phosphates,sulfates, and carbamates (e.g., N,N-dimethylaminocarbonyl).

A specific example of a monoterpene that may be used in the presentinvention is perillyl alcohol. The derivatives of perillyl alcoholinclude perillyl alcohol carbamates, perillyl alcohol esters, perillicaldehydes, dihydroperillic acid, perillic acid, perillic aldehydederivatives, dihydroperillic acid esters and perillic acid esters.

In certain embodiments, a POH carbamate is synthesized by a processcomprising the step of reacting a first reactant of perillylchloroformate with a second reactant such as dimethyl celocoxib (DMC),temozolomide (TMZ) and rolipram. The reaction may be carried out in thepresence of tetrahydrofuran and a base such as n-butyl lithium. Perillylchloroformate may be made by reacting POH with phosgene. For example,POH conjugated with temozolomide through a carbamate bond may besynthesized by reacting temozolomide with oxalyl chloride followed byreaction with perillyl alcohol. The reaction may be carried out in thepresence of 1,2-dichloroethane.

As described herein, the monoterpene derivative, such as POH carbamateconjugated with another therapeutic agent, can be further reacted with afatty acid to form a tri-conjugate structure containing an acylatedaliphatic tail (which is the fatty acid with the terminal —OH of thecarboxyl group removed). The fatty acid can be unsaturated, such asCaprylic acid, Capric acid, Lauric acid, Myristic acid, Palmitic acid,Stearic acid, Arachidic acid, Behenic acid, Lignoceric acid, Ceroticacid; or saturated, such as Myristoleic acid, Palmitoleic acid, Sapienicacid, Oleic acid, Elaidic acid, Vaccenic acid, Linoleic acid,Linoelaidic acid, α-Linolenic acid, Arachidonic acid, Eicosapentaenoicacid, Erucic acid, Docosahexaenoic acid. For example, when thetherapeutic agent is TMZ, the fatty acid can be linoleic acid to form(3-Methyl-4-oxo-3,4-dihydro-imidazo[5,1-d][1,2,3,5]tetrazine-8-carbonyl)-octadeca-9,12-dienoyl-carbamicacid 4-isopropenyl-cyclohex-1-enylmethyl ester, or it can be palmiticacid to form(3-Methyl-4-oxo-3,4-dihydro-imidazo[5,1-d][1,2,3,5]tetrazine-8-carbonyl)-hexadecanoicacid 4-isopropenyl-cyclohex-1-enylmethyl ester. Halides of the fattyacid (e.g., acyl bromide, acyl chloride of these fatty acids), oranhydrides of the fatty acid can also be used. For example, a POHcarbamate that is a conjugate with a therapeutic agent can be furtherreacted with a fatty acid (or a fatty acid halide or anhydride) suchthat the hydrogen of the —NH— group in the carbamate linker group issubstituted with the acylated aliphatic tail of the fatty acid.

Except where stated otherwise, the following definitions applythroughout the present specification and claims. These definitions applyregardless of whether a term is used by itself or in combination withother terms. For example, the definition of “alkyl” applies not only toalkyl groups per se, but also to the alkyl portions of alkoxy,alkylamino, alkylthio or alkylcarbonyl groups etc. Furthermore allranges described for a chemical group, for example “from 1 to 13 carbonatoms” or “C₁-C₆ alkyl” include all combinations and sub-combinations ofranges and specific numbers of carbon atoms therein.

As described herein, “Alkyl” means a straight (linear) chain or branchedchain aliphatic hydrocarbon group having from 4 to 28 carbon atoms inthe chain. Preferred alkyl groups have from 10 to 20 carbon atoms in thechain. More preferred alkyl groups have from 14 to 20 carbon atoms inthe chain. Non limiting examples of suitable alkyl groups includeisopropyl, sec-butyl, n-butyl, and t-butyl.

As described herein “Alkenyl” means a straight (linear) chain orbranched chain aliphatic hydrocarbon group having 1 to 8, preferably 1to 5, more preferably 1 to 3, carbon-carbon double bonds and having from4 to 28 carbon atoms in the chain. Preferred alkenyl groups have from 10to 20 carbon atoms in the chain. More preferred alkenyl groups have from14 to 20 carbon atoms in the chain. Non limiting examples of suitablealkenyl groups include isopropenyl, n-butenyl, 1-hexenyl and3-methylbut-2-enyl.

As described herein “Alkynyl” means a straight (linear) chain orbranched chain aliphatic hydrocarbon group having at 1 to 5carbon-carbon triple bonds and having from 4 to 28 carbon atoms in thechain. Preferred alkynyl groups have from 4 to 12 carbon atoms in thechain. More preferred alkynyl groups have from 4 to 6 carbon atoms inthe chain. Examples of suitable 2-propynyl and 2-butynyl.

According to the present invention, the therapeutic agents that may beconjugated with monoterpene (or sesquiterpene) include, but are notlimited to, chemotherapeutic agents, therapeutic agents for treatment ofCNS disorders (including, without limitation, primary degenerativeneurological disorders such as Alzheimer's, Parkinson's, multiplesclerosis, Attention-Deficit Hyperactivity Disorder or ADHD,psychological disorders, psychosis and depression), immunotherapeuticagents, angiogenesis inhibitors, and anti-hypertensive agents.Anti-cancer agents that may be conjugated with monoterpene orsesquiterpene can have one or more of the following effects on cancercells or the subject: cell death; decreased cell proliferation;decreased numbers of cells; inhibition of cell growth; apoptosis;necrosis; mitotic catastrophe; cell cycle arrest; decreased cell size;decreased cell division; decreased cell survival; decreased cellmetabolism; markers of cell damage or cytotoxicity; indirect indicatorsof cell damage or cytotoxicity such as tumor shrinkage; improvedsurvival of a subject; or disappearance of markers associated withundesirable, unwanted, or aberrant cell proliferation. U.S. PatentPublication No. 20080275057.

Also encompassed by the present invention is admixtures and/orcoformulations of a monoterpene (or sesquiterpene) and at least onetherapeutic agent.

Chemotherapeutic agents include, but are not limited to, DNA alkylatingagents, topoisomerase inhibitors, endoplasmic reticulum stress inducingagents, a platinum compound, an antimetabolite, vincalkaloids, taxanes,epothilones, enzyme inhibitors, receptor antagonists, tyrosine kinaseinhibitors, boron radiosensitizers (i.e. velcade), and chemotherapeuticcombination therapies.

Non-limiting examples of DNA alkylating agents are nitrogen mustards,such as Cyclophosphamide (Ifosfamide, Trofosfamide), Chlorambucil(Melphalan, Prednimustine), Bendamustine, Uramustine and Estramustine;nitrosoureas, such as Carmustine (BCNU), Lomustine (Semustine),Fotemustine, Nimustine, Ranimustine and Streptozocin; alkyl sulfonates,such as Busulfan (Mannosulfan, Treosulfan); Aziridines, such asCarboquone, Triaziquone, Triethylenemelamine; Hydrazines (Procarbazine);Triazenes such as Dacarbazine and Temozolomide (TMZ); Altretamine andMitobronitol.

Non-limiting examples of Topoisomerase I inhibitors include Campothecinderivatives including SN-38, APC, NPC, campothecin, topotecan, exatecanmesylate, 9-nitrocamptothecin, 9-aminocamptothecin, lurtotecan,rubitecan, silatecan, gimatecan, diflomotecan, extatecan, BN-80927,DX-8951f, and MAG-CPT as described in Pommier Y. (2006) Nat. Rev. Cancer6(10):789-802 and U.S. Patent Publication No. 200510250854;Protoberberine alkaloids and derivatives thereof including berberrubineand coralyne as described in Li et al. (2000) Biochemistry39(24):7107-7116 and Gatto et al. (1996) Cancer Res. 15(12):2795-2800;Phenanthroline derivatives including Benzo[i]phenanthridine, Nitidine,and fagaronine as described in Makhey et al. (2003) Bioorg. Med. Chem.11 (8): 1809-1820; Terbenzimidazole and derivatives thereof as describedin Xu (1998) Biochemistry 37(10):3558-3566; and Anthracyclinederivatives including Doxorubicin, Daunorubicin, and Mitoxantrone asdescribed in Foglesong et al. (1992) Cancer Chemother. Pharmacol.30(2):123-]25, Crow et al. (1994) J. Med. Chem. 37(19):31913194, andCrespi et al. (1986) Biochem. Biophys. Res. Commun. 136(2):521-8.Topoisomerase II inhibitors include, but are not limited to Etoposideand Teniposide. Dual topoisomerase I and II inhibitors include, but arenot limited to, Saintopin and other Naphthecenediones, DACA and otherAcridine-4-Carboxamindes, Intoplicine and other Benzopyridoindoles,TAS-I03 and other 7H-indeno[2,1-c]Quinoline-7-ones, Pyrazoloacridine, XR11576 and other Benzophenazines, XR 5944 and other Dimeric compounds,7-oxo-7H-dibenz[f,ij]Isoquinolines and 7-oxo-7H-benzo[e]pyrimidines, andAnthracenyl-amino Acid Conjugates as described in Denny and Baguley(2003) Curr. Top. Med. Chem. 3(3):339-353. Some agents inhibitTopoisomerase II and have DNA intercalation activity such as, but notlimited to, Anthracyclines (Aclarubicin, Daunorubicin, Doxorubicin,Epirubicin, Idarubicin, Amrubicin, Pirarubicin, Valrubicin, Zorubicin)and Antracenediones (Mitoxantrone and Pixantrone).

Examples of endoplasmic reticulum stress inducing agents include, butare not limited to, dimethyl-celecoxib (DMC), nelfinavir, celecoxib, andboron radiosensitizers (i.e. velcade (Bortezomib)).

Platinum based compounds are a subclass of DNA alkylating agents.Non-limiting examples of such agents include Cisplatin, Nedaplatin,Oxaliplatin, Triplatin tetranitrate, Satraplatin, Aroplatin, Lobaplatin,and JM-216. (see McKeage et al. (1997) J. Clin. Oncol. 201:1232-1237 andin general, CHEMOTHERAPY FOR GYNECOLOGICAL NEOPLASM, CURRENT THERAPY ANDNOVEL APPROACHES, in the Series Basic and Clinical Oncology, Angioli etal. Eds., 2004).

“FOLFOX” is an abbreviation for a type of combination therapy that isused to treat colorectal cancer. It includes 5-FU, oxaliplatin andleucovorin. Information regarding this treatment is available on theNational Cancer Institute's web site, cancer.gov, last accessed on Jan.16, 2008.

“FOLFOX/BV” is an abbreviation for a type of combination therapy that isused to treat colorectal cancer. This therapy includes 5-FU,oxaliplatin, leucovorin and Bevacizumab. Furthermore, “XELOX/BV” isanother combination therapy used to treat colorectal cancer, whichincludes the prodrug to 5-FU, known as Capecitabine (Xeloda) incombination with oxaliplatin and bevacizumab. Information regardingthese treatments are available on the National Cancer Institute's website, cancer.gov or from 23 the National Comprehensive Cancer Network'sweb site, nccn.org, last accessed on May 27, 2008.

Non-limiting examples of antimetabolite agents include Folic acid based,i.e. dihydrofolate reductase inhibitors, such as Aminopterin,Methotrexate and Pemetrexed; thymidylate synthase inhibitors, such asRaltitrexed, Pemetrexed; Purine based, i.e. an adenosine deaminaseinhibitor, such as Pentostatin, a thiopurine, such as Thioguanine andMercaptopurine, a halogenated/ribonucleotide reductase inhibitor, suchas Cladribine, Clofarabine, Fludarabine, or a guanine/guanosine:thiopurine, such as Thioguanine; or Pyrimidine based, i.e.cytosine/cytidine: hypomethylating agent, such as Azacitidine andDecitabine, a DNA polymerase inhibitor, such as Cytarabine, aribonucleotide reductase inhibitor, such as Gemcitabine, or athymine/thymidine: thymidylate synthase inhibitor, such as aFluorouracil (5-FU). Equivalents to 5-FU include prodrugs, analogs andderivative thereof such as 5′-deoxy-5-fluorouridine (doxifluroidine),1-tetrahydrofuranyl-5-fluorouracil (ftorafur), Capecitabine (Xeloda),S-I (MBMS-247616, consisting of tegafur and two modulators, a5-chloro-2,4-dihydroxypyridine and potassium oxonate), ralititrexed(tomudex), nolatrexed (Thymitaq, AG337), LY231514 and ZD9331, asdescribed for example in Papamicheal (1999) The Oncologist 4:478-487.

Examples of vincalkaloids, include, but are not limited to Vinblastine,Vincristine, Vinflunine, Vindesine and Vinorelbine.

Examples of taxanes include, but are not limited to docetaxel,Larotaxel, Ortataxel, Paclitaxel and Tesetaxel. An example of anepothilone is iabepilone.

Examples of enzyme inhibitors include, but are not limited tofarnesyltransferase inhibitors (Tipifarnib); CDK inhibitor (Alvocidib,Seliciclib); proteasome inhibitor (Bortezomib); phosphodiesteraseinhibitor (Anagrelide; rolipram); IMP dehydrogenase inhibitor(Tiazofurine); and lipoxygenase inhibitor (Masoprocol). Examples ofreceptor antagonists include, but are not limited to ERA (Atrasentan);retinoid X receptor (Bexarotene); and a sex steroid (Testolactone).

Examples of tyrosine kinase inhibitors include, but are not limited toinhibitors to ErbB: HER1/EGFR (Erlotinib, Gefitinib, Lapatinib,Vandetanib, Sunitinib, Neratinib); HER2/neu (Lapatinib, Neratinib); RTKclass III: C-kit (Axitinib, Sunitinib, Sorafenib), FLT3 (Lestaurtinib),PDGFR (Axitinib, Sunitinib, Sorafenib); and VEGFR (Vandetanib,Semaxanib, Cediranib, Axitinib, Sorafenib); bcr-ab1 (Imatinib,Nilotinib, Dasatinib); Src (Bosutinib) and Janus kinase 2(Lestaurtinib).

“Lapatinib” (Tykerb®) is an dual EGFR and erbB-2 inhibitor. Lapatinibhas been investigated as an anticancer monotherapy, as well as incombination with trastuzumab, capecitabine, letrozole, paclitaxel andFOLFIRI (irinotecan, 5-fluorouracil and leucovorin), in a number ofclinical trials. It is currently in phase III testing for the oraltreatment of metastatic breast, head and neck, lung, gastric, renal andbladder cancer.

A chemical equivalent of lapatinib is a small molecule or compound thatis a tyrosine kinase inhibitor (TKI) or alternatively a HER-1 inhibitoror a HER-2 inhibitor. Several TKIs have been found to have effectiveantitumor activity and have been approved or are in clinical trials.Examples of such include, but are not limited to, Zactima (ZD6474),Iressa (gefitinib), imatinib mesylate (STI571; Gleevec), erlotinib(OSI-1774; Tarceva), canertinib (CI 1033), semaxinib (SU5416), vatalanib(PTK787/ZK222584), sorafenib (BAY 43-9006), sutent (SUI 1248) andlefltmomide (SU101).

PTK/ZK is a tyrosine kinase inhibitor with broad specificity thattargets all VEGF receptors (VEGFR), the platelet-derived growth factor(PDGF) receptor, c-KIT and c-Fms. Drevs (2003) Idrugs 6(8):787-794.PTK/ZK is a targeted drug that blocks angiogenesis and lymphangiogenesisby inhibiting the activity of all known receptors that bind VEGFincluding VEGFR-I (Flt-1), VEGFR-2 (KDR/Flk-1) and VEGFR-3 (Flt-4). Thechemical names of PTK/ZK are1-[4-Chloroanilino]-4-[4-pyridylmethyl]phthalazine Succinate or1-Phthalazinamine, N-(4-chlorophenyl)-4-(4-pyridinylmethyl)-butanedioate(1:1). Synonyms and analogs of PTK/TK are known as Vatalanib, CGP79787D,PTK787/ZK 222584, CGP-79787, DE-00268, PTK-787, PTK787A, VEGFR-TKinhibitor, ZK 222584 and ZK.

Chemotherapeutic agents that can be conjugated with monoterpene orsesquiterpene may also include amsacrine, Trabectedin, retinoids(Alitretinoin, Tretinoin), Arsenic trioxide, asparagine depleterAsparaginase/Pegaspargase), Celecoxib, Demecolcine, Elesclomol,Elsamitrucin, Etoglucid, Lonidamine, Lucanthone, Mitoguazone, Mitotane,Oblimersen, Temsirolimus, and Vorinostat.

The monoterpene or sesquiterpene derivative may be conjugated withangiogenesis inhibitors. Examples of angiogenesis inhibitors include,but are not limited to, angiostatin, angiozyme, antithrombin III,AG3340, VEGF inhibitors, batimastat, bevacizumab (avastin), BMS-275291,CAI, 2C3, HuMV833 Canstatin, Captopril, carboxyamidotriazole, cartilagederived inhibitor (CDI), CC-5013,6-O-(chloroacetyl-carbonyl)-fumagillol, COL-3, combretastatin,combretastatin A4 Phosphate, Dalteparin, EMD 121974 (Cilengitide),endostatin, erlotinib, gefitinib (Iressa), genistein, halofuginonehydrobromide, Idl, Id3, IM862, imatinib mesylate, IMC-IC11 Inducibleprotein 10, interferon-alpha, interleukin 12, lavendustin A, LY317615 orAE-941, marimastat, mspin, medroxpregesterone acetate, Meth-1, Meth-2,2-methoxyestradiol (2-ME), neovastat, oteopontin cleaved product, PEX,pigment epithelium growth factor (PEGF), platelet factor 4, prolactinfragment, proliferin-related protein (PRP), PTK787/ZK 222584, ZD6474,recombinant human platelet factor 4 (rPF4), restin, squalamine, SU5416,SU6668, SU11248 suramin, Taxol, Tecogalan, thalidomide, thrombospondin,TNP-470, troponin-1, vasostatin, VEG1, VEGF-Trap, and ZD6474.

Non-limiting examples of angiogenesis inhibitors also include, tyrosinekinase inhibitors, such as inhibitors of the tyrosine kinase receptorsFlt-1 (VEGFR1) and Flk-1/KDR (VEGFR2), inhibitors of epidermal-derived,fibroblast-derived, or platelet derived growth factors, MMP (matrixmetalloprotease) inhibitors, integrin blockers, pentosan polysulfate,angiotensin II antagonists, cyclooxygenase inhibitors (includingnon-steroidal anti-inflammatory drugs (NSAIDs) such as aspirin andibuprofen, as well as selective cyclooxygenase-2 inhibitors such ascelecoxib and rofecoxib), and steroidal anti-inflammatories (such ascorticosteroids, mineralocorticoids, dexamethasone, prednisone,prednisolone, methylpred, betamethasone).

Other therapeutic agents that modulate or inhibit angiogenesis and mayalso be conjugated with monoterpene or sesquiterpene include agents thatmodulate or inhibit the coagulation and fibrinolysis systems, including,but not limited to, heparin, low molecular weight heparins andcarboxypeptidase U inhibitors (also known as inhibitors of activethrombin activatable fibrinolysis inhibitor [TAFIa]). U.S. PatentPublication No. 20090328239. U.S. Pat. No. 7,638,549.

Non-limiting examples of the anti-hypertensive agents includeangiotensin converting enzyme inhibitors (e.g., captopril, enalapril,delapril etc.), angiotensin II antagonists (e.g., candesartan cilexetil,candesartan, losartan (or Cozaar), losartan potassium, eprosartan,valsartan (or Diovan), termisartan, irbesartan, tasosartan, olmesartan,olmesartan medoxomil etc.), calcium antagonists (e.g., manidipine,nifedipine, amlodipine (or Amlodin), efonidipine, nicardipine etc.),diuretics, renin inhibitor (e.g., aliskiren etc.), aldosteroneantagonists (e.g., spironolactone, eplerenone etc.), beta-blockers(e.g., metoprolol (or Toporol), atenolol, propranolol, carvedilol,pindolol etc.), vasodilators (e.g., nitrate, soluble guanylate cyclasestimulator or activator, prostacycline etc.), angiotensin vaccine,clonidine and the like. U.S. Patent Publication No. 20100113780.

Other therapeutic agents that may be conjugated with monoterpene (orsesquiterpene) include, but are not limited to, Sertraline (Zoloft),Topiramate (Topamax), Duloxetine (Cymbalta), Sumatriptan (Imitrex),Pregabalin (Lyrica), Lamotrigine (Lamictal), Valaciclovir (Valtrex),Tamsulosin (Flomax), Zidovudine (Combivir), Lamivudine (Combivir),Efavirenz (Sustiva), Abacavir (Epzicom), Lopinavir (Kaletra),Pioglitazone (Actos), Desloratidine (Clarinex), Cetirizine (Zyrtec),Pentoprazole (Protonix), Lansoprazole (Prevacid), Rebeprazole (Aciphex),Moxifloxacin (Avelox), Meloxicam (Mobic), Dorzolamide (Truspot),Diclofenac (Voltaren), Enlapril (Vasotec), Montelukast (Singulair),Sildenafil (Viagra), Carvedilol (Coreg), Ramipril (Delix).

Table 1 lists pharmaceutical agents that can be conjugated withmonoterpene (or sesquiterpene), including structure of thepharmaceutical agent and the preferred derivative for conjugation.

TABLE 1 Brand Generic Preferred Name Name Activity Structure DerivativeZoloft Sertraline Depression

Carbamate Topamax Topiramate Seizures

Carbamate Cymbalta Duloxetine Depression

Carbamate Imitrex Sumatriptan Migraine

Carbamate Lyrica Pregabalin Neuropathic pain

Carbamate or Ester Lamictal Lamotrigine Seizures

Carbamate Valtrex Valaciclovir Herpes

Carbamate Tarceva Erlotinib Non-small cell lung cancer

Carbamate Flomax Tamsulosin Benign prostatic Cancer

Carbamate Gleevec Imatinib Leukemia

Carbamate Combivir Zidovudine HIV infection

Carbamate Combivir Lamivudine HIV infection

Carbonate Sustiva                         Epzicom Efavirenz                        Abacavir HIV infection                         HIV infection

Carbamate                         Carbamate Kaletra Lopinavir HIVinfection

Carbamate Actos Pioglitazone Type-2 diabetes

Carbamate Clarinex Desloratidine Allergic rhinitis

Carbamate Zyrtec Cetirizine Allergic

Ester Protonix Pentoprazole Gastrointestinal

Carbamate Prevacid Lansoprazole Gastrointestinal

Carbamate Aciphex Rebeprazole Gastrointestinal

Carbamate Diovan Valsartan Hypertension

Carbamate Cozaar Losartan Hypertension

Carbamate Avelox Moxifloxacin Bacterial infection

Carbamate or Ester Mobic                   Truspot Meloxicam                  Dorzolamide Osteoarthritis                   Intraocular pressure

Carbamate                   Carbamate Voltaren Diclofenac Osteoarthritis& rheumatoid arthritis

Carbamate or Ester Vasotec Enlapril Hypertension

Carbamate or Ester Singulair Montelukast Asthma

Ester Amlodin Amlodipine Hypertension

Carbamate Toporol Metoprolol Hypertension

Carbamate Viagra Sildenafil Erectile dysfunction

Carbamate Coreg Carvedilol Hypertension

Carbamate Delix                                     Sinemet (Parcopa,Atamet) Ramipril                                     L-DOPA Hypertension                                    Neurological disorders

Carbamate or Ester

The purity of the monoterpene (or sesquiterpene) derivatives may beassayed by gas chromatography (GC) or high pressure liquidchromatography (HPLC). Other techniques for assaying the purity ofmonoterpene (or sesquiterpene) derivatives and for determining thepresence of impurities include, but are not limited to, nuclear magneticresonance (NMR) spectroscopy, mass spectrometry (MS), GC-MS, infraredspectroscopy (IR), and thin layer chromatography (TLC). Chiral puritycan be assessed by chiral GC or measurement of optical rotation.

The monoterpene (or sesquiterpene) derivatives may be purified bymethods such as crystallization, or by separating the monoterpene (orsesquiterpene) derivative from impurities according to the uniquephysicochemical properties (e.g., solubility or polarity) of thederivative. Accordingly, the monoterpene (or sesquiterpene) derivativecan be separated from the monoterpene (or sesquiterpene) by suitableseparation techniques known in the art, such as preparativechromatography, (fractional) distillation, or (fractional)crystallization.

The invention also provides for methods of using the herein describedmonoterpenes (or sesquiterpenes) derivatives to treat a disease, such ascancer or other nervous system disorders. For example, the cancer is atumor of the nervous system, e.g., glioblastoma. The cancer can also bea skin cancer, such as melanoma, basal cell carcinoma, or squamous cellcarcinoma. The disease can also be a precancerous skin lesion.

A monoterpenes (or sesquiterpenes) derivative may be administered alone,or in combination with radiation, surgery or additional chemotherapeuticagents. A monoterpene or sesquiterpene derivative may also beco-administered with antiviral agents, anti-inflammatory agents orantibiotics. The agents may be administered concurrently orsequentially. A monoterpene (or sesquiterpene) derivative can beadministered before, during or after the administration of the otheractive agent(s).

The monoterpene or sesquiterpene derivative may be used in combinationwith radiation therapy. In one embodiment, the present inventionprovides for a method of treating tumor cells, such as malignant gliomacells, with radiation, where the cells are treated with an effectiveamount of a monoterpene derivative, such as a perillyl alcoholcarbamate, and then exposed to radiation. Monoterpene derivativetreatment may be before, during and/or after radiation. For example, themonoterpene or sesquiterpene derivative may be administered continuouslybeginning one week prior to the initiation of radiotherapy and continuedfor two weeks after the completion of radiotherapy. U.S. Pat. Nos.5,587,402 and 5,602,184.

In one embodiment, the present invention provides for a method oftreating tumor cells, such as malignant glioma cells, with chemotherapy,where the cells are treated with an effective amount of a monoterpenederivative, and then exposed to chemotherapy. Monoterpene derivativetreatment may be before, during and/or after chemotherapy.

The monoterpene (or sesquiterpene) derivatives of the present inventionmay be used for the treatment of nervous system cancers, such as amalignant glioma (e.g., astrocytoma, anaplastic astrocytoma,glioblastoma multiforme), retinoblastoma, pilocytic astrocytomas (gradeI), meningiomas, metastatic brain tumors, neuroblastoma, pituitaryadenomas, skull base meningiomas, and skull base cancer. As used herein,the term “nervous system tumors” refers to a condition in which asubject has a malignant proliferation of nervous system cells.

Cancers that can be treated by the present monoterpene (orsesquiterpene) derivatives include, but are not limited to, lung cancer,ear, nose and throat cancer, leukemia, colon cancer, melanoma,pancreatic cancer, mammary cancer, prostate cancer, breast cancer,hematopoietic cancer, ovarian cancer, basal cell carcinoma, biliarytract cancer; bladder cancer; bone cancer; breast cancer; cervicalcancer; choriocarcinoma; colon and rectum cancer; connective tissuecancer; cancer of the digestive system; endometrial cancer; esophagealcancer; eye cancer; cancer of the head and neck; gastric cancer;intra-epithelial neoplasm; kidney cancer; larynx cancer; leukemiaincluding acute myeloid leukemia, acute lymphoid leukemia, chronicmyeloid leukemia, chronic lymphoid leukemia; liver cancer; lymphomaincluding Hodgkin's and Non-Hodgkin's lymphoma; myeloma; fibroma,neuroblastoma; oral cavity cancer (e.g., lip, tongue, mouth, andpharynx); ovarian cancer; pancreatic cancer; prostate cancer;retinoblastoma; rhabdomyosarcoma; rectal cancer; renal cancer; cancer ofthe respiratory system; sarcoma; skin cancer; stomach cancer; testicularcancer; thyroid cancer; uterine cancer; cancer of the urinary system, aswell as other carcinomas and sarcomas. U.S. Pat. No. 7,601,355.

The present invention also provides methods of treating CNS disorders,including, without limitation, primary degenerative neurologicaldisorders such as Alzheimer's, Parkinson's, psychological disorders,psychosis and depression. Treatment may consist of the use of amonoterpene or sesquiterpene derivative alone or in combination withcurrent medications used in the treatment of Parkinson's, Alzheimer's,or psychological disorders.

The present invention also provides a method of improvingimmunomodulatory therapy responses comprising the steps of exposingcells to an effective amount of a monoterpene or sisquiterpenederivative, such as a POH-TMZ-fatty acid derivative, before or duringimmunomodulatory treatment. Preferred immunomodulatory agents arecytokines, such interleukins, lymphokines, monokines, interfereons andchemokines.

The present composition may be administered by any method known in theart, including, without limitation, intranasal, oral, transdermal,ocular, intraperitoneal, inhalation, intravenous, ICV, intracisternalinjection or infusion, subcutaneous, implant, vaginal, sublingual,urethral (e.g., urethral suppository), subcutaneous, intramuscular,intravenous, rectal, sub-lingual, mucosal, ophthalmic, spinal,intrathecal, intra-articular, intra-arterial, sub-arachinoid, bronchialand lymphatic administration. Topical formulation may be in the form ofgel, ointment, cream, aerosol, etc; intranasal formulation can bedelivered as a spray or in a drop; transdermal formulation may beadministered via a transdermal patch or iontorphoresis; inhalationformulation can be delivered using a nebulizer or similar device.Compositions can also take the form of tablets, pills, capsules,semisolids, powders, sustained release formulations, solutions,suspensions, elixirs, aerosols, or any other appropriate compositions.

To prepare such pharmaceutical compositions, one or more of monoterpene(or sesquiterpene) derivatives may be mixed with a pharmaceuticalacceptable carrier, adjuvant and/or excipient, according to conventionalpharmaceutical compounding techniques. Pharmaceutically acceptablecarriers that can be used in the present compositions encompass any ofthe standard pharmaceutical carriers, such as a phosphate bufferedsaline solution, water, and emulsions, such as an oil/water or water/oilemulsion, and various types of wetting agents. The compositions canadditionally contain solid pharmaceutical excipients such as starch,cellulose, talc, glucose, lactose, sucrose, gelatin, malt, rice, flour,chalk, silica gel, magnesium stearate, sodium stearate, glycerolmonostearate, sodium chloride, dried skim milk and the like. Liquid andsemisolid excipients may be selected from glycerol, propylene glycol,water, ethanol and various oils, including those of petroleum, animal,vegetable or synthetic origin, e.g., peanut oil, soybean oil, mineraloil, sesame oil, etc. Liquid carriers, particularly for injectablesolutions, include water, saline, aqueous dextrose, and glycols. Forexamples of carriers, stabilizers and adjuvants, see Remington'sPharmaceutical Sciences, edited by E. W. Martin (Mack PublishingCompany, 18th ed., 1990). The compositions also can include stabilizersand preservatives.

As used herein, the term “therapeutically effective amount” is an amountsufficient to treat a specified disorder or disease or alternatively toobtain a pharmacological response treating a disorder or disease.Methods of determining the most effective means and dosage ofadministration can vary with the composition used for therapy, thepurpose of the therapy, the target cell being treated, and the subjectbeing treated. Treatment dosages generally may be titrated to optimizesafety and efficacy. Single or multiple administrations can be carriedout with the dose level and pattern being selected by the treatingphysician. Suitable dosage formulations and methods of administering theagents can be readily determined by those of skill in the art. Forexample, the composition are administered at about 0.01 mg/kg to about200 mg/kg, about 0.1 mg/kg to about 100 mg/kg, or about 0.5 mg/kg toabout 50 mg/kg. When the compounds described herein are co-administeredwith another agent or therapy, the effective amount may be less thanwhen the agent is used alone.

Transdermal formulations may be prepared by incorporating the activeagent in a thixotropic or gelatinous carrier such as a cellulosicmedium, e.g., methyl cellulose or hydroxyethyl cellulose, with theresulting formulation then being packed in a transdermal device adaptedto be secured in dermal contact with the skin of a wearer. If thecomposition is in the form of a gel, the composition may be rubbed ontoa membrane of the patient, for example, the skin, preferably intact,clean, and dry skin, of the shoulder or upper arm and or the uppertorso, and maintained thereon for a period of time sufficient fordelivery of the monoterpene (or sesquiterpene) derivative to the bloodserum of the patient. The composition of the present invention in gelform may be contained in a tube, a sachet, or a metered pump. Such atube or sachet may contain one unit dose, or more than one unit dose, ofthe composition. A metered pump may be capable of dispensing one metereddose of the composition.

The present invention also provides the compositions as described abovefor intranasal administration. As such, the compositions can furthercomprise a permeation enhancer. Southall et al. Developments in NasalDrug Delivery, 2000. The monoterpene (or sesquiterpene) derivative maybe administered intranasally in a liquid form such as a solution, anemulsion, a suspension, drops, or in a solid form such as a powder, gel,or ointment. Devices to deliver intranasal medications are well known inthe art. Nasal drug delivery can be carried out using devices including,but not limited to, intranasal inhalers, intranasal spray devices,atomizers, nasal spray bottles, unit dose containers, pumps, droppers,squeeze bottles, nebulizers, metered dose inhalers (MDI), pressurizeddose inhalers, insufflators, and bi-directional devices. The nasaldelivery device can be metered to administer an accurate effectivedosage amount to the nasal cavity. The nasal delivery device can be forsingle unit delivery or multiple unit delivery. In a specific example,the ViaNase Electronic Atomizer from Kurve Technology (Bethell, Wash.)can be used in this invention (http://www.kurvetech.com). The compoundsof the present invention may also be delivered through a tube, acatheter, a syringe, a packtail, a pledget, a nasal tampon or bysubmucosal infusion. U.S. Patent Publication Nos. 20090326275,20090291894, 20090281522 and 20090317377.

The monoterpene (or sesquiterpene) derivative can be formulated asaerosols using standard procedures. The monoterpene (or sesquiterpene)derivative may be formulated with or without solvents, and formulatedwith or without carriers. The formulation may be a solution, or may bean aqueous emulsion with one or more surfactants. For example, anaerosol spray may be generated from pressurized container with asuitable propellant such as, dichlorodifluoromethane,trichlorofluoromethane, dichlorotetrafluoroethane, hydrocarbons,compressed air, nitrogen, carbon dioxide, or other suitable gas. Thedosage unit can be determined by providing a valve to deliver a meteredamount. Pump spray dispensers can dispense a metered dose or a dosehaving a specific particle or droplet size. As used herein, the term“aerosol” refers to a suspension of fine solid particles or liquidsolution droplets in a gas. Specifically, aerosol includes a gas-bornesuspension of droplets of a monoterpene (or sesquiterpene), as may beproduced in any suitable device, such as an MDI, a nebulizer, or a mistsprayer. Aerosol also includes a dry powder composition of thecomposition of the instant invention suspended in air or other carriergas. Gonda (1990) Critical Reviews in Therapeutic Drug Carrier Systems6:273-313. Raeburn et al., (1992) Pharmacol. Toxicol. Methods27:143-159.

The monoterpene (or sesquiterpene) derivative may be delivered to thenasal cavity as a powder in a form such as microspheres delivered by anasal insufflator. The monoterpene (or sesquiterpene) derivative may beabsorbed to a solid surface, for example, a carrier. The powder ormicrospheres may be administered in a dry, air-dispensable form. Thepowder or microspheres may be stored in a container of the insufflator.Alternatively the powder or microspheres may be filled into a capsule,such as a gelatin capsule, or other single dose unit adapted for nasaladministration.

The pharmaceutical composition can be delivered to the nasal cavity bydirect placement of the composition in the nasal cavity, for example, inthe form of a gel, an ointment, a nasal emulsion, a lotion, a cream, anasal tampon, a dropper, or a bioadhesive strip. In certain embodiments,it can be desirable to prolong the residence time of the pharmaceuticalcomposition in the nasal cavity, for example, to enhance absorption.Thus, the pharmaceutical composition can optionally be formulated with abioadhesive polymer, a gum (e.g., xanthan gum), chitosan (e.g., highlypurified cationic polysaccharide), pectin (or any carbohydrate thatthickens like a gel or emulsifies when applied to nasal mucosa), amicrosphere (e.g., starch, albumin, dextran, cyclodextrin), gelatin, aliposome, carbamer, polyvinyl alcohol, alginate, acacia, chitosansand/or cellulose (e.g., methyl or propyl; hydroxyl or carboxy;carboxymethyl or hydroxylpropyl).

The composition containing the purified monoterpene (or sesquiterpene)can be administered by oral inhalation into the respiratory tract, i.e.,the lungs.

Typical delivery systems for inhalable agents include nebulizerinhalers, dry powder inhalers (DPI), and metered-dose inhalers (MDI).

Nebulizer devices produce a stream of high velocity air that causes atherapeutic agent in the form of liquid to spray as a mist. Thetherapeutic agent is formulated in a liquid form such as a solution or asuspension of particles of suitable size. In one embodiment, theparticles are micronized. The term “micronized” is defined as havingabout 90% or more of the particles with a diameter of less than about 10μm. Suitable nebulizer devices are provided commercially, for example,by PARI GmbH (Starnberg, Germany). Other nebulizer devices includeRespimat (Boehringer Ingelheim) and those disclosed in, for example,U.S. Pat. Nos. 7,568,480 and 6,123,068, and WO 97/12687. Themonoterpenes (or sesquiterpenes) can be formulated for use in anebulizer device as an aqueous solution or as a liquid suspension.

DPI devices typically administer a therapeutic agent in the form of afree flowing powder that can be dispersed in a patient's air-streamduring inspiration. DPI devices which use an external energy source mayalso be used in the present invention. In order to achieve a freeflowing powder, the therapeutic agent can be formulated with a suitableexcipient (e.g., lactose). A dry powder formulation can be made, forexample, by combining dry lactose having a particle size between about 1μm and 100 μm with micronized particles of the monoterpenes (orsesquiterpenes) and dry blending. Alternatively, the monoterpene can beformulated without excipients. The formulation is loaded into a drypowder dispenser, or into inhalation cartridges or capsules for use witha dry powder delivery device. Examples of DPI devices providedcommercially include Diskhaler (GlaxoSmithKline, Research Triangle Park,N.C.) (see, e.g., U.S. Pat. No. 5,035,237); Diskus (GlaxoSmithKline)(see, e.g., U.S. Pat. No. 6,378,519; Turbuhaler (AstraZeneca,Wilmington, Del.) (see, e.g., U.S. Pat. No. 4,524,769); and Rotahaler(GlaxoSmithKline) (see, e.g., U.S. Pat. No. 4,353,365). Further examplesof suitable DPI devices are described in U.S. Pat. Nos. 5,415,162,5,239,993, and 5,715,810 and references therein.

MDI devices typically discharge a measured amount of therapeutic agentusing compressed propellant gas. Formulations for MDI administrationinclude a solution or suspension of active ingredient in a liquefiedpropellant. Examples of propellants include hydrofluoroalklanes (HFA),such as 1,1,1,2-tetrafluoroethane (HFA 134a) and1,1,1,2,3,3,3-heptafluoro-n-propane, (HFA 227), and chlorofluorocarbons,such as CCl₃F. Additional components of HFA formulations for MDIadministration include co-solvents, such as ethanol, pentane, water; andsurfactants, such as sorbitan trioleate, oleic acid, lecithin, andglycerin. (See, for example, U.S. Pat. No. 5,225,183, EP 0717987, and WO92/22286). The formulation is loaded into an aerosol canister, whichforms a portion of an MDI device. Examples of MDI devices developedspecifically for use with HFA propellants are provided in U.S. Pat. Nos.6,006,745 and 6,143,227. For examples of processes of preparing suitableformulations and devices suitable for inhalation dosing see U.S. Pat.Nos. 6,268,533, 5,983,956, 5,874,063, and 6,221,398, and WO 99/53901, WO00/61108, WO 99/55319 and WO 00/30614.

The monoterpene (or sesquiterpene) derivative may be encapsulated inliposomes or microcapsules for delivery via inhalation. A liposome is avesicle composed of a lipid bilayer membrane and an aqueous interior.The lipid membrane may be made of phospholipids, examples of whichinclude phosphatidylcholine such as lecithin and lysolecithin; acidicphospholipids such as phosphatidylserine and phosphatidylglycerol; andsphingophospholipids such as phosphatidylethanolamine and sphingomyelin.Alternatively, cholesterol may be added. A microcapsule is a particlecoated with a coating material. For example, the coating material mayconsist of a mixture of a film-forming polymer, a hydrophobicplasticizer, a surface activating agent or/and a lubricantnitrogen-containing polymer. U.S. Pat. Nos. 6,313,176 and 7,563,768.

The monoterpene (or sesquiterpene) derivative may also be used alone orin combination with other chemotherapeutic agents via topicalapplication for the treatment of localized cancers such as breast canceror melanomas. The monoterpene (or sesquiterpene) derivative may also beused in combination with narcotics or analgesics for transdermaldelivery of pain medication.

This invention also provides the compositions as described above forocular administration. As such, the compositions can further comprise apermeation enhancer. For ocular administration, the compositionsdescribed herein can be formulated as a solution, emulsion, suspension,etc. A variety of vehicles suitable for administering compounds to theeye are known in the art. Specific non-limiting examples are describedin U.S. Pat. Nos. 6,261,547; 6,197,934; 6,056,950; 5,800,807; 5,776,445;5,698,219; 5,521,222; 5,403,841; 5,077,033; 4,882,150; and 4,738,851.

The monoterpene (or sesquiterpene) derivative can be given alone or incombination with other drugs for the treatment of the above diseases fora short or prolonged period of time. The present compositions can beadministered to a mammal, preferably a human. Mammals include, but arenot limited to, murines, rats, rabbit, simians, bovines, ovine, porcine,canines, feline, farm animals, sport animals, pets, equine, andprimates.

The present invention also provides a method for inhibiting the growthof a cell in vitro, ex vivo or in vivo, where a cell, such as a cancercell, is contacted with an effective amount of the monoterpene (orsesquiterpene) derivative as described herein.

Pathological cells or tissue such as hyperproliferative cells or tissuemay be treated by contacting the cells or tissue with an effectiveamount of a composition of this invention. The cells, such as cancercells, can be primary cancer cells or can be cultured cells availablefrom tissue banks such as the American Type Culture Collection (ATCC).The pathological cells can be cells of a systemic cancer, gliomas,meningiomas, pituitary adenomas, or a CNS metastasis from a systemiccancer, lung cancer, prostate cancer, breast cancer, hematopoieticcancer or ovarian cancer. The cells can be from a vertebrate, preferablya mammal, more preferably a human. U.S. Patent Publication No.2004/0087651. Balassiano et al. (2002) Intern. J. Mol. Med. 10:785-788.Thorne, et al. (2004) Neuroscience 127:481-496. Fernandes, et al. (2005)Oncology Reports 13:943-947. Da Fonseca, et al. (2008) SurgicalNeurology 70:259267. Da Fonseca, et al. (2008) Arch. Immunol. Ther. Exp.56:267-276. Hashizume, et al. (2008) Neuroncology 10:112-120.

In vitro efficacy of the present composition can be determined usingmethods well known in the art. For example, the cytoxicity of thepresent monoterpene (or sesquiterpene) and/or the therapeutic agents maybe studied by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazoliumbromide] cytotoxicity assay. MTT assay is based on the principle ofuptake of MTT, a tetrazolium salt, by metabolically active cells whereit is metabolized into a blue colored formazon product, which can beread spectrometrically. J. of Immunological Methods 65: 55 63, 1983. Thecytoxicity of the present monoterpene (or sesquiterpene) derivativeand/or the therapeutic agents may be studied by colony formation assay.Functional assays for inhibition of VEGF secretion and IL-8 secretionmay be performed via ELISA. Cell cycle block by the present monoterpene(or sesquiterpene) derivative and/or the therapeutic agents may bestudied by standard propidium iodide (PI) staining and flow cytometry.Invasion inhibition may be studied by Boyden chambers. In this assay alayer of reconstituted basement membrane, Matrigel, is coated ontochemotaxis filters and acts as a barrier to the migration of cells inthe Boyden chambers. Only cells with invasive capacity can cross theMatrigel barrier. Other assays include, but are not limited to cellviability assays, apoptosis assays, and morphological assays.

The following are examples of the present invention and are not to beconstrued as limiting.

EXAMPLES Example 1 Preparation of POH-TMZ-linoleate (orTMZ-POH-linoleate, TPL) triconjugate((3-Methyl-4-oxo-3,4-dihydro-imidazo[5,1-d][1,2,3,5]tetrazine-8-carbonyl)-octadeca-9,12-dienoyl-carbamic acid4-isopropenyl-cyclohex-1-enylmethyl ester)

The reaction scheme is as follows:

Oxalyl chloride (0.64 g, 5.04 mmol) was added slowly to a mixture ofTemozolamide (Source: OChem Incorporation, Lot #9110918A; 0.5 g, 2.57mmol) in 1,2-dichloroethane (10 mL) over a period of 2 min whilemaintaining the temperature at 10° C. under N₂. The reaction mixture wasallowed to warm to room temperature and then heated to reflux for 3 h.The excess of oxalyl chloride and 1,2-dichloroethane were removed byconcentration under vacuum. The resulting residue was redissolved in1,2-dichlorethane (15 mL) and the reaction mixture was cooled to 10° C.under N₂. A solution of Perillyl alcohol (0.086 g, 0.56 mmol) in1,2-dichloroethane (3 mL) was added over a period of 5 min. The reactionmixture was allowed to warm to room temperature and stirred for 14 h.1,2-Dichloroethane was concentrated under vacuum to give a residue whichwas triturated with hexanes. The resulting pale-yellow solid (TMZ-POH orPOH-TMZ) was filtered and washed with hexanes. Weight: 0.85 g; Yield:89%. ¹H-NMR (400 MHz, CDCl₃): δ 1.4-2.2 (m, 10H), 4.06 (s, 3H), 4.6-4.8(m, 4H), 5.88 (br s, 1H), 8.42 (s, 1H), 9.31 (br s, 1H); MS (APCI): Nomolecular ion peak is observed. m/e: 314 (100%), 286.5 (17%), 136 (12%).

POH-TMZ obtained above (300 mg, 0.80 mmol) in dry THF (5.0 mL) was addedto Sodium hydride (60%, 48 mg, 1.2 mmol) in dry THF (4.0 mL) at 0-5° C.The mixture was allowed to warm to 20-25° C. and stirred for 1.0 h.Linoleoyl chloride (alternatively the acid chloride of any fatty acid orcarboxylic acid, R—COCl, may be used) solution (264 mg, 0.88 mmol) indry THF was added slowly over a period of 15 min, while maintaining thetemperature below 10° C. The mixture was slowly heated to 35-40° C. andstirred for 1.0 h. After confirming the completion of the reaction byTLC (20% EtOAc/Hexanes), the mixture was quenched with saturatedammonium chloride solution. The reaction mixture was extracted withethyl acetate (2×15 mL). The combined organic layer was washed withwater (25 mL), brine (10%, 25 mL) and dried over sodium sulfate. Thefiltered organic layer was concentrated to give an oil which waspurified by column chromatography [Column dimensions: dia: 1.5 cm,height: 10 cm, silica: 230-400 mesh] and eluted with a mixture of 5%ethyl acetate/hexanes (100 mL) followed by 10% ethyl acetate/hexanes(100 mL). The 10% ethyl acetate/hexanes fractions were combined andconcentrated under vacuum to give a gummy solid. Weight: 260 mg; Yield:50%.

Example 2 Cytotoxicity of POH-TMZ-Linoleate (TPL)

The cytotoxicity of TMZ, POH-TMZ, and POH-TMZ-Linoleate were studied bycolony formation assay of A2058 and A375 melanoma cells. A2058 and A375cells were harvested and made single cell suspension in culture medium(DMEM with 10% FBS, 1% Penicillin/Streptomycin). 500 cells/well wereseeded into 12-well plate (tissue culture treated, from OlympusPlastics) pre-warmed culture medium at 37° C. The cells were culturedovernight for attachment to the plate. Then the medium was removed andreplaced with fresh medium containing varied concentrations of drugs (orno drugs as a control) for further 48-hour incubation. After 48 hourstreatment, the medium was aspirated and replaced with normal culturemedium. 7-10 days later, the culture medium was aspirated, and thecolonies formed inside the wells were washed with cold PBS once. Thenthe colonies were stained with 0.1% methylene blue (in methanol) for 4-6hours. The plates were then washed and the formed colonies counted (thecolonies formed with 20 cells or more are countable). The colonies countresults shown in FIGS. 1 and 2 indicate that, at similar concentrations,POH-TMZ-linoleate was more effective in killing the melanoma cells thanPOH-TMZ, which is in turn more effective than TMZ.

Example 3 In Vivo Tumor Growth Inhibition by POH-TMZ-Linoleate (TPL)

Inhibition of tumor growth by POH-TMZ-linoleate was studied in nudemice. Nude mice were injected subcutaneously with 2×10⁶ human melanomatumor cells, A2058. The melanoma cells were allowed to form a palpablenodule in the mice. When the tumor size reached 1.0-1.5 cm in diameterin any dimension, a control mouse was treated topically with a vehicle(10% DMSO in 45% Glycerol+45% ethanol), and a test mouse was topicallytreated with TPL (50 mM TPL working solution was reconstituted in 45%Glycerol+45% Ethanol, and 100 μl (25 mg/kg) was applied topically on theskin which covers the whole tumor). The control mouse and the test mousewere both treated twice daily for 14 days. It was observed that thetumor of the test mouse developed a hard surface after 14 days oftreatment (FIG. 3B), with reduced vascularization compared with thetumor before the treatment (FIG. 3A).

After the 14 days of treatment the mice were sacrificed, the tumorremoved, sectioned at 8 μm, fixed with acetone, and stored at −20° C.For histological analysis, the tissues were blocked with sea block andstained with the primary antibody Rat anti Mouse (1:50) overnightfollowing which secondary biotinylated goat anti rat antibody (1:200)was added for 45 minutes, washed and avidin biotin complex, ABC elitewas added for 30 minutes, washed and then stained with AEC and counterstained with hematoxylin. Such staining was used to evaluate theendothelial marker CD31 expression, which is representative of theamounts of blood vessels in the tumor. Staining images were captured at40× under optical microscope. A visual inspection of the stained tissuesshowed that the tumor angiogenesis was much lower in the TPL-treatedmouse (FIGS. 4D-4F) than that in the vehicle-treated mouse (FIGS.4A-4C). The histological analysis results were further quantified usingImageJ software by measuring the average area of staining in the tumorsof the test mouse after TPL treatment and the control mouse, where theaverage area of staining was calculated based on the red precipitate ofthe stain in μm². From FIG. 5, it can be seen that the average area ofstaining is much smaller in the tumor of the test mouse than that in thecontrol mouse. These comparisons demonstrate the efficacy of TPL intreating melanoma.

The scope of the present invention is not limited by what has beenspecifically shown and described hereinabove. Variations, modificationsand other implementations of what is described herein will occur tothose of ordinary skill in the art without departing from the spirit andscope of the invention.

What is claimed is:
 1. A compound of the Formula I;

wherein R is selected from the group consisting of a C₄ to C₂₈ linear orbranched alkyl, linear or branched C₄ to C₂₈ alkeneyl and a linear orbranched C₄ to C₂₈ alkynyl group; or a pharmaceutically acceptable saltthereof.
 2. The compound according to claim 1 wherein R is a C₄ to C₂₈linear or branched alkeneyl containing 1, 2, 3, 4 or 5 double bonds or alinear or branched C₄ to C₂₈ alkynyl group containing 1, 2, 3, 4 or 5triple bonds.
 3. The compound according to claim 1 of the Formula II;

or a pharmaceutically acceptable salt thereof.
 4. A method of treatingmelanoma in a mammal, comprising the step of administering to the mammala therapeutically effective amount of the compound according to claim 1,2 or
 3. 5. The method of claim 4, wherein the compound is administeredtopically.
 6. A pharmaceutical composition comprising a compoundaccording to claim 1, 2 or
 3. 7. A compound of the Formula III;

wherein R₁ is selected from a C₄ to C₂₈ linear or branched alkyl, linearor branched C₄ to C₂₈ alkeneyl or a linear or branched C₄ to C₂₈ alkynylgroup; or a pharmaceutically acceptable salt thereof.
 8. The compoundaccording to claim 7 wherein R₁ is a C₄ to C₂₈ linear or branchedalkeneyl containing 1, 2, 3, 4 or 5 double bonds or a linear or branchedC₄ to C₂₈ alkynyl group containing 1, 2, 3, 4 or 5 triple bonds.
 9. Thecompound according to claim 7 of the Formula IV; or a pharmaceuticallyacceptable salt thereof


10. A pharmaceutical composition comprising a compound according toclaim 7, 8 or
 9. 11. A process for making a compound of the Formula I

comprising the steps of a) reacting Temozolamide with oxalyl chloride ina halogenated solvent to give the isocyanate of Temozolamide Formula A

b) reacting the isocyanate of Temozolamide with perillyl alcohol in ahalogenated solvent to afford a compound of Formula B

c) reacting the compound of Formula B with an acid chloride of theFormula C

wherein R is selected from the group consisting of a C₄ to C₂₈ linear orbranched alkyl, linear or branched C₄ to C₂₈ alkeneyl containing 1, 2,3, 4, or 5 double bonds or a linear or branched C₄ to C₂₈ alkynyl group;containing 1, 2, 3, 4 or 5 triple bonds in the presence of a base in anethereal solvent to afford a compound of the Formula I.
 12. The processaccording to claim 11 wherein the halogenated solvent is 1,2dichloroethane.
 13. The process according to claim 11 wherein the acidchloride is linoleic acid chloride.
 14. The process according to claim11 wherein the base is NaH.
 15. The process according to claim 11wherein the ethereal solvent is THF.